Persistent growth effects of temperature and photoperiod in Atlantic cod Gadus morhua

Short-term environmental manipulations during the early juvenile stage have a large impact on harvesting size of Atlantic cod Gadus morhua nearly 3 years later. A group of juvenile Atlantic cod (initial mass 9·5 g) were reared for 3 months under simulated natural photoperiod or continuous light, and a range of temperatures (7, 10, 13 and 16° C, and a group called T-step, i.e. with temperature reduced successively from 16 to 13 and 10° C). After termination of the laboratory trial, the fish were moved to sea pens and reared at ambient conditions for 30 months before harvesting in June 2006. Observed growth gain from the 3 month laboratory trial was still persistent following the 30 months of sea-pen ongrowing. The T-step group displayed 15, 13, 1 and 10% superior mass gain respectively than the groups initially at 7, 10, 13 and 16° C at harvest in June 2006. Similarly, rearing under continuous light during the initial 3 month period during the early juvenile stage resulted in 1–9% larger size at harvesting compared to fish reared at simulated natural photoperiod. Gonado-somatic and hepato-somatic indices were similar in all groups. Contribution to the understanding of the mechanism behind size variation in adult fish can have wide range applications for Atlantic cod fisheries and aquaculture.     

Restricted T cell receptor V-β and J-β usage in T cells from interleukin-2-cultured lymphocytes of ovarian and renal carcinomas

Tumour-infiltrating lymphocytes (TIL) are often observed in human tumours and their presence has been correlated with a better prognosis. It has been suggested that TIL are enriched for tumour-specific cytotoxic cells, and TIL activated and expanded in vitro by interleukin-2 (IL-2) are currently used in the therapy of human cancer. We have studied the T cell repertoire in IL-2-expanded TIL cells from patients with ovarian and renal carcinoma using T-cell-receptor-V--specific monoclonal antibodies and a polymerase-chain-reaction-based Southern blot technique for analysis of J- usage. In TIL lines derived from three of nine patients with ovarian carcinomas and from two of eight patients with renal carcinomas, selective usage of the V-6 or V-5 T-cell receptor gene products was found. The majority of the cells were CD4⁺, with up to 40% of the T cells utilizing the same V- gene. T-cell lines derived from peripheral blood lymphocytes from patients or healthy donors contained normal levels of V- subsets. Only moderate levels of V-6⁺ T cells were detected from freshly isolated TIL and the increase of this subpopulation appeared as a result of in vitro culture. The level of clonal restriction, as measured by the usage of J- gene segments within the V-5 or V-6 families, was analysed using a recently developed technique based on the polymerase chain reaction. Evidence for restricted J- usage was detected only in TIL expanded in vitro, while this was not the case in freshly isolated tumour-derived lymphocytes or T cell lines obtained from peripheral blood lymphocytes. The presence of a population with biased T cell receptor expression in cells derived from tumour tissue could be explained by their activation in vivo as a result of contact with tumour antigens and should be taken into consideration when discussing the therapeutic efficiency of IL-2-expanded TIL.     

Serological cloning of cancer/testis antigens expressed in prostate cancer using cDNA phage surface display

Serological cloning of tumor-associated antigens (TAAs) using patient autoantibodies and tumor cDNA expression libraries (SEREX) has identified a wide array of tumor proteins eliciting B-cell responses in patients. However, alternative cloning strategies with the possibility of high throughput analysis of patient sera and tumor libraries may be of interest. We explored the pJuFo phage surface display system, allowing display of recombinant tumor proteins on the surface of M13 filamentous phage, for cloning of TAAs in prostate cancer (PC). Control experiments established that after a few rounds of selection on immobilized specific IgG, a high degree of enrichment of seroreactive clones was achieved. With an increasing number of selection rounds, a higher yield of positive clones was offset by an apparent loss of diversity in the repertoire of selected clones. Using autologous patient serum IgG in a combined biopanning and immunoscreening approach, we identified 13 different TAAs. Three of these (NY-ESO-1, Lage-1, and Xage-1) were known members of the cancer/testis family of TAAs, and one other protein had previously been isolated by SEREX in cancer types other than PC. Specific IgG responses against NY-ESO-1 were found in sera from 4/20 patients with hormone refractory PC, against Lage-1 in 3/20, and Xage-1 in 1/20. No reactivity against the remaining proteins was detected in other PC patients, and none of the TAAs reacted with serum from healthy subjects. The results demonstrate that phage surface display combined with postselection immunoscreening is suitable for cloning a diverse repertoire of TAAs from tumor tissue cDNA libraries. Furthermore, candidate TAAs for vaccine development of PC were identified.     

Enhancement of streptolysin O activity and intrinsic cytotoxic effects of the group A streptococcal toxin, NAD-glycohydrolase

Streptolysin O (SLO) is a cholesterol-dependent cytolysin produced by the important human pathogen, group A Streptococcus (Streptococcus pyogenes or GAS). In addition to its cytolytic activity, SLO mediates the translocation of GAS NAD-glycohydrolase (NADase) into human epithelial cells in vitro. Production of both NADase and SLO is associated with augmented host cell injury beyond that produced by SLO alone, but the mechanism of enhanced cytotoxicity is not known. We have now shown that expression of NADase together with SLO dramatically enhanced the lytic activity of GAS culture supernatants for erythrocytes but had no effect on SLO-mediated poration of synthetic cholesterol-rich liposomes. This result revealed a previously unknown contribution of NADase to the cytolytic activity associated with GAS production of SLO. Purified recombinant SLO bound NADase in vitro, supporting a specific, physical interaction of the two proteins. Exposure of human keratinocytes to wild-type GAS, but not to a NADase-deficient mutant strain, resulted in profound depletion of cellular NAD+ and ATP. Furthermore, expression of recombinant GAS NADase in yeast, in the absence of SLO, induced growth arrest, depletion of NAD+ and ATP, and cell death. These findings have provided evidence that the augmentation of SLO-mediated cytotoxicity by NADase is a consequence of depletion of host cell energy stores through the enzymatic action of NADase. Together, the results have provided mechanistic insight into the cytotoxic effects of a unique bipartite bacterial toxin.     

Carotenoid and protein supplementation have differential effects on pheasant ornamentation and immunity

A currently popular hypothesis states that the expression of carotenoid-dependent sexual ornaments and immune function may be correlated because both traits are positively affected by carotenoids. However, such a correlation may arise for another reason: it is well known that immune function is dependent on nutritional condition. A recent study has suggested that the expression of ornaments may too depend on nutritional condition, as males in good nutritional condition are better at assimilating and/or modulating carotenoids. Thus, carotenoid-dependent ornaments and immune function may be correlated because both are dependent on nutritional condition. To elucidate if, and how, ornamentation and immune function are linked, pheasant diets were supplemented with carotenoid and/or protein in a fully factorial experiment. Carotenoid treatment affected wattle coloration and tail growth, but not cellular or humoral immunity. Immunity was unrelated to males' initial ornamentation including wattle colour. Males in better body condition, measured as residual mass, increased their wattle coloration more when carotenoid supplemented. Protein positively affected humoral but not cellular immunity, but had no effect on ornaments. Cellular, but not humoral, immunity increased with male body condition. Thus, there was no evidence that an immune-stimulatory effect of carotenoids resulted in wattle coloration honestly signalling immune function, but wattle coloration may still signal male body condition.     

A new hypothesis to explain allocation of dry matter between mycorrhizal fungi and pine seedlings in relation to nutrient supply

Nutrient uptake by forest trees is largely dependent on their associated ectomycorrhizal fungi. The presence of extramatrical mycelium produced by ectomycorrhizal fungi allows trees to exploit a larger soil volume. In this paper the effects of macronutrients on the production of extramatrical mycelium are reviewed. It is concluded that elevated levels of nitrogen and, to some extent, phosphorus strongly inhibit the development of extramatrical mycelium. A deficiency of phosphorus, on the other hand, stimulates ectomycorrhizal development. Low levels of phosphorus may offset the negative influence of nitrogen, indicating that the nitrogen effect is indirect. No other macronutrients have been shown to affect extramatrical mycelium significantly, however, very few studies have been made.To explain reduced ectomycorrhizal development under conditions of high N availability, it has been suggested that the host would allocate less carbohydrate to the mycobiont under such conditions owing to a greater demand for carbon by growing shoots. In the present paper an alternative explanation is suggested: The fungus is forced to take up all available nitrogen and must therefore consume the available carbohydrate in order to assimilate it. The surplus of carbohydrates after nitrogen assimilation can then be used to produce fungal mycelium and fruit bodies. However, the total allocation of host carbohydrate to the mycorrhizal fungus is not reduced at elevated levels of N supply. In contrast with previous theories, the present one proposes that it is the fungus, rather than the host which adjusts its carbon allocation patterns to the N supply.     

Identification of acylation products in SHAPE chemistry

SHAPE chemistry (selective 2′-hydroxyl acylation analyzed by primer extension) has been developed to specifically target flexible nucleotides (often unpaired nucleotides) independently to their purine or pyrimidine nature for RNA secondary structure determination. However, to the best of our knowledge, the structure of 2′-O-acylation products has never been confirmed by NMR or X-ray data. We have realized the acylation reactions between cNMP and NMIA under SHAPE chemistry conditions and identified the acylation products using standard NMR spectroscopy and LC–MS/MS experiments. For cAMP and cGMP, the major acylation product is the 2′-O-acylated compound (>99%). A trace amount of N-acylated cAMP has also been identified by LC–UV–MS². While for cCMP, the isolated acylation products are composed of 96% of 2′-O-acylated, 4% of N,O-diacylated, and trace amount of N-acylated compounds. In addition, the characterization of the major 2′-O-acylated compound by NMR showed slight differences in the conformation of the acylated sugar between the three cyclic nucleotides. This interesting result should be useful to explain some unexpected reactivity of the SHAPE chemistry.     

Photosynthetic formation of inorganic pyrophosphate in phototrophic bacteria

In this paper we report studies on photosynthetic formation of inorganic pyrophosphate (PP_i) in three phototrophic bacteria. Formation of PP_i was found in chromatophores from Rhodopseudomonas viridis but not in chromatophores from Rhodopseudomonas blastica and Rhodobacter capsulatus. The maximal rate of PP_i synthesis in Rps. viridis was 0.15 mol PP_i formed/(min*mol Bacteriochlorophyll) at 23°C. The synthesis of PP_i was inhibited by electron transport inhibitors, uncouplers and fluoride, but was insensitive to oligomycin and venturicidin. The steady state rate of PP_i synthesis under continuous illumination was about 15% of the steady-state rate of ATP synthesis. The synthesis of PP_i after short light flashes was also studied. The yield of PP_i after a single 1 ms flash was equivalent to approximately 1 mol PP_i/500 mol Bacteriochlorophyll. In Rps. viridis chromatophores, PP_i was also found to induce a membrane potential, which was sensitive to carbonyl cyanide p-trifluoromethoxyphenylhydrazone and NaF.Abbreviations BChl Bacteriochlorophyll - F₀F₁-ATPase Membrane bound proton translocating ATP synthase - FCCP Carbonyl cyanide p-trifluoromethoxyphenylhydrazone - H⁺-PPase Membrane bound proton translocating PP_i synthase - TPP⁺ Tetraphenyl phosphonium ion - TPB^- Tetraphenyl boron ion - Transmembrane electrical potential difference     

Meta-analysis identified the TNFA -308G > A promoter polymorphism as a risk factor for disease severity in patients with rheumatoid arthritis

Introduction

The goal of this study is to investigate whether the -308G > A promoter polymorphism in the tumor necrosis factor alpha (TNFA) gene is associated with disease severity and radiologic joint damage in a large cohort of patients with rheumatoid arthritis (RA).

Methods

A long-term observational early RA inception cohort (n = 208) with detailed information about disease activity and radiologic damage after 3, 6 and 9 years of disease was genotyped for the TNFA -308G > A promoter polymorphism (rs1800629). A longitudinal regression analysis was performed to assess the effect of genotype on RA disease severity and joint damage. Subsequently, a meta-analysis, including all publically available data, was performed to further test the association between joint erosions and the TNFA polymorphism. To learn more about the mechanism behind the effect of the polymorphism, RNA isolated from peripheral blood from RA patients (n = 66) was used for TNFA gene expression analysis by quantitative PCR.

Results

Longitudinal regression analysis with correction for gender and disease activity showed a significant difference in total joint damage between GG and GA+AA genotype groups (P = 0.002), which was stable over time. The meta-analysis, which included 2,053 patients, confirmed an association of the genetic variant with the development of erosions (odds ratio 0.78, 95% CI 0.62, 0.98). No significant differences in TNFA gene expression were observed for the different genotypes, confirming earlier findings in healthy individuals.

Conclusions

Our data confirm that the TNFA -308G > A promoter polymorphism is associated with joint damage in patients with RA. This is not mediated by differences in TNFA gene expression between genotypes.